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User Manual H201-NIKON-TI-SR June 2015 H201-NIKON-TI-SR Compatible with the following XY stages Nikon T/Ti-SR H201-T-UNIT-BL Page 1 Compatible with the following Okolab Controllers www.oko-lab.com H201-NIKON-TI-SR User Manual June 2015 Index 1. Components and dimensions................................................................................................... 3 2. Sample Holders ................................................................................................................... 3 2.1 Available Sample Holders....................................................................................................... 3 3. Available Lids ..................................................................................................................... 4 3.1 Sliding lid .......................................................................................................................... 5 3.2 Koehler Lid ........................................................................................................................ 6 3.3 Injection Lid ...................................................................................................................... 6 3.4 Laser Interlock Lid ............................................................................................................... 7 4. Insertion of the Sample Feedback Temperature Sensor .................................................................. 8 5. Insertion of Sample Holder into Chamber ................................................................................... 8 6. Working with 35 or 60 mm Petri Dish – Spacing Rings and Magnetic Locks ........................................... 9 7. Working with 1x3’’ and 1x2’’ chamber slides - magnetic locks ........................................................11 8. Connecting the Gas Supply and Filling the Humidity Module ...........................................................12 9. Working with Perfusion ........................................................................................................12 Page 2 10. Connecting the Chamber with XY stage ....................................................................................13 www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 1. Components and dimensions H201-NIKON-TI-SR includes the following components: Chamber main body Sliding glass lid for easy pipetting Integrated humidity module. It is not necessary to select an additional humidity module Sliding Lid Distance between focal plane and upper side of the glass lid Sample Holder-Order Separately Chamber top view Integrated Humidty Module Chamber Base Figure 1. H201-NIKON-TI-SR - Components and Dimensions. 2. Sample Holders 2.1 Available Sample Holders The following sample holders are available. NOTE: Please contact [email protected] if you cannot find the sample holder you are looking for. We are constantly adding new inserts to the list. #1 35mm Petri-dish 1xGS-M single #1 1x3in. chamber slide 1xLABTEK-M single #1 Lab-Tek 1in.x2in. chambered cover glass 1xLABTEK-II-M single #1 Lab-Tek II 1in.x2in. chambered cover glass 1x60-M single #1 60mm Petri-dish Page 3 1x35-M single www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 1x35-M single For #1 35mm Petri-dish 1x60-M single For #1 60mm Petri-dish 1xLABTEK-M single For #1 Lab-Tek 1’’x2’’ chambered cover glass 1xGS-M single For #1 1’’x3’’ chamber slide 1xLABTEK-II-M single For #1 Lab-Tek II 1’’x2’’ chambered cover glass Figure 2.Available sample holders. 3. Available Lids The following glass Lids are available for H201-NIKON-TI-SR : H201-SLIDING-LID-2: For easy sample loading and pipetting – Included in code H201-NIKON-TI-SR H201-KOEHLER-LID-2*: Reduces chamber height to 22 mm and allows imaging under Koehler illumination H201-INJECTION-LID-2*: Glass lid with two small openings (sealed with flexible plastic) allowing injection or permanent access to the sample. Compatible only with sample holder 1x35-M single H201-LASER-INTERLOCK-LID-2*: Glass lid with safety switch. Connects to laser controller and automatically turns laser off when lid is lifted. Compatible only with sample holder 1x35-M single Page 4 * OPTIONAL – not included with H201-NIKON-TI-SR www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 3.1 Sliding lid The Sliding Lid is a glass lid allowing for easy sample loading and pipetting. The Sliding Lid is screwed onto the chamber. Figure 3 shows chamber dimensions with the Sliding Lid. Figure 4 illustrates how to remove Sliding Lid, when a different Lid is necessary. NOTE: Sliding Lid MUST BE REMOVED when using any other Lid. The Sliding Lid is fixed onto the chamber with 4 screws. Screws location is indicated by letter A in Figure 4 (image 2 and 3). Keep Sliding Lid fully closed to access screws labeled A in image 2. Open Sliding Lid to access screws labeled A in image 3. Sliding Lid Distance between focal plane and upper side of the glass lid Chamber top view Figure 3. H201-NIKON-TI-SR (Sliding Lid comes as standard) Screws M2x6 Sliding Lid A A 1 2 3 Page 5 Figure 4. Assembly of the sliding lid www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 3.2 Koehler Lid The Koehler Lid is a glass lid reducing the chamber’s height to 22 mm and allowing imaging under Koehler illumination. The Koehler Lid is placed onto the chamber (NO SCREWS or TOOLS are necessary). Figure 5 shows chamber dimensions with Koehler lid. H201-KOHLER LID-2 Distance between focal plane and upper side of the glass lid Chamber top view Figure 5. H201-NIKON-TI-SR + H201-KOEHLER-LID-2 3.3 Injection Lid The Injection Lid is a glass lid with two small openings sealed with flexible plastic allowing injection or permanent access to the sample. NOTE: Compatible with sample holder 1x35-M single ONLY. The Injection Lid is placed onto the chamber (NO SCREWS or TOOLS are necessary). Page 6 Figure 6 shows chamber dimensions with the Injection Lid. www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 H201-INJECTION LID-2 Distance between focal plane and upper side of the glass lid Chamber top view Figure 6. H201-NIKON-TI-SR + H201-INJECTION-LID 3.4 Laser Interlock Lid The Laser Interlock Lid is a glass lid with a safety switch connecting to the laser controller. It automatically turns the laser off when the lid is lifted. NOTE: Compatible with sample holder 1x35-M single ONLY. The Laser Interlock Lid is placed onto the chamber (NO SCREWS or TOOLS are necessary). Figure 7 shows chamber dimensions with the Laser Interlock Lid. H201-LASER-INTERLOCK LID-2 Distance between focal plane and upper side of the glass lid Page Figure 7. H201-NIKON-TI-SR + H201-LASER-INTERLOCK-LID 7 Chamber top view www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 4. Insertion of the Sample Feedback Temperature Sensor Insert the Sample Feedback Temperature Sensor through the dedicated opening located in the H201-NIKON-TI-SR (see Figure 8, Frontal and 3D views). Temperature Sensor dedicated opening Temperature Sensor dedicated opening 1.Frontal view 2.3D view Figure 8. Insertion of the temperature sensor inside the chamber. 5. Insertion of Sample Holder into Chamber Sample holders fit into the chamber base and are held in place by magnets embedded within both chamber and holder. To introduce a sample holder with the proper orientation, match the red dot on the holder to the one on the chamber base, as illustrated in Figure 9. Insert the adapter by superimposing the red points Page 8 Figure 9. Introduction of the Specimen Holder inside the Chamber Base. www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 6. Working with 35 or 60 mm Petri Dish – Spacing Rings and Magnetic Locks Magnetic locks prevent movement of 35 and 60 mm dishes inside the sample holder. Figure 10 illustrates the available magnetic locks for 35 and 60 mm dishes. Threaded magnetic posts allow adjusting holder’s height. Magnetic Locks for 35 and 60 mm petri dishes. Included in the corresponding sample holder Figure 10.Magnetic locks for 35 and 60 mm dish NOTE: Magnetic locks are included with sample holder. Spacing rings with variable diameter are also included in according to the needs of the user. Figure 11 shows how to position the ring inside the sample holder. Ring selection guide diagrams: rings for 35 and 60 mm dish -Figure 12 and Figure 13, respectively. 1 2 3 Figure 11. 1) Insertion of the ring, 2) insertion of the 35 mm dish, 3) insertion of the magnetic lock. Page 9 NOTE: Spacing rings are included with sample holder. www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 BD Falcon-35x10 Willco-35x10 Grainer-Petri35x10 Ibidi μ-Dish35mm-low Corning-35x10mm MatTek-P35G-X-14-X Figure 12. Ring selection scheme for 35 mm dish. Ibidi μ-dish 50mm-low MatTek-P50G-X-30-X BD Falcon 60x15 Grainer - Petri 60x15 Corning-60x15mm Willco60x15 Page 10 Figure 13. Ring selection scheme for 60 mm dish. www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 7. Working with 1x3’’ and 1x2’’ chamber slides - magnetic locks Magnetic locks prevent movement of 1’’x 3’’ and 1’’x 2’’ chamber slides inside of the sample holder. NOTE: Magnetic locks are included with sample holder. 2 1 Figure 14. Magnetic lock for 1'' x 3'' chamber slide. 3 1 2 Page 11 Figure 15. Magnetic lock for 1'' x 2'' chamber slide. To lock the slide, push simultaneously the buttons indicated with (3). www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 8. Connecting the Gas Supply and Filling the Humidity Module Single silicon tubing carries output gas from the Okolab Gas Controller to the H201-NIKON-TI-SR. Silicon tubing connects to a gas input - brass opening - located on a side of the H201-NIKON-TI-SR. See Figure 16. Connect the silicon tubing by gently pushing it onto brass opening. H201-NIKON-TI-SR includes an integrated humidity module in order to maintain a high humidity percentage inside the chamber. Fill the humidity module inside the chamber through the water input - brass opening - located on a side of the H201-NIKON-TI-SR (See Figure 16) using a silicon tubing 2 mm ID and a syringe*. *NOTE: Do not exceed 0.09 l Gas Input Water Input Figure 16. Connection with gas supply. 9. Working with Perfusion H201-NIKON-TI-SR features 8 perfusion holes for the insertion of perfusion tubing up to 2.5 mm in outer diameter. Small screws plug the perfusion holes when not in use. (Grub screws M3x3). Remove small screws as needed before introducing perfusion tubing. Figure 17 shows location of perfusion holes. Perfusion dedicated opening 1. Frontal view Page Figure 17. Perfusion 12 2. 3D view www.oko-lab.com User Manual H201-NIKON-TI-SR June 2015 10. Connecting the Chamber with XY stage Follow the steps shown in the images Figure 18 and listed below in order to correctly connect the chamber with XY stage. 1. Place the chamber on the stage and tighten 4 captive screws (See Figure 18 Image 1 and Image 2). Captive screws housings are indicated with letter B in Figure 18 Image 2 2. Use a 2.5mm metric Allen Wrench to tighten the four captive screws while keeping the chamber sliding lid open (See Figure 18 Image 2) Captive Screws A 1 B 2 Page Figure 18. Connection of the Chamber with XY Stage 13 3 www.oko-lab.com